scholarly journals Analysis of Genes Induced by Sendai Virus Infection of Mutant Cell Lines Reveals Essential Roles of Interferon Regulatory Factor 3, NF-κB, and Interferon but Not Toll-Like Receptor 3

2005 ◽  
Vol 79 (7) ◽  
pp. 3920-3929 ◽  
Author(s):  
Christopher P. Elco ◽  
Jeanna M. Guenther ◽  
Bryan R. G. Williams ◽  
Ganes C. Sen
Keyword(s):  
Virus Infection ◽  
Cell Lines ◽  
Sendai Virus ◽  
Mutant Cell ◽  
Gene Induction ◽  
Interferon Regulatory Factor ◽  
Toll Like Receptor ◽  
Regulatory Factor ◽  
Interferon Regulatory Factor 3 ◽  
Infected Cells

ABSTRACT Sendai virus (SeV) infection causes the transcriptional induction of many cellular genes that are also induced by interferon (IFN) or double-stranded RNA (dsRNA). We took advantage of various mutant cell lines to investigate the putative roles of the components of the IFN and dsRNA signaling pathways in the induction of those genes by SeV. Profiling the patterns of gene expression in SeV-infected cells demonstrated that Toll-like receptor 3, although essential for gene induction by dsRNA, was dispensable for gene induction by SeV. In contrast, Jak1, which mediates IFN signaling, was required for the induction of a small subset of genes by SeV. NF-κB and interferon regulatory factor 3 (IRF-3), the two major transcription factors activated by virus infection, were essential for the induction of two sets of genes by SeV. As expected, some of the IRF-3-dependent genes, such as ISG56, were more strongly induced by SeV in IRF-3-overexpressing cells. Surprisingly, in those cells, a number of NF-κB-dependent genes, such as the A20 gene, were induced poorly. Using a series of cell lines expressing increasing levels of IRF-3, we demonstrated that the degree of induction of A20 mRNA, upon SeV infection, was inversely proportional to the cellular level of IRF-3, whereas that of ISG56 mRNA was directly proportional. Thus, IRF-3 can suppress the expression of NF-κB-dependent genes in SeV-infected cells.

scholarly journals Hsp90 Regulates Activation of Interferon Regulatory Factor 3 and TBK-1 Stabilization in Sendai Virus-infected Cells

2006 ◽  
Vol 17 (3) ◽  
pp. 1461-1471 ◽  
Author(s):  
Kai Yang ◽  
Hexin Shi ◽  
Rong Qi ◽  
Shaogang Sun ◽  
Yujie Tang ◽  
...  
Keyword(s):  
Sendai Virus ◽  
Specific Interaction ◽  
Hybrid Approach ◽  
Specific Inhibitor ◽  
Interferon Regulatory Factor ◽  
Dominant Negative ◽  
Regulatory Factor ◽  
Interferon Regulatory Factor 3 ◽  
Infected Cells ◽  
Irf3 Activation

Interferon regulatory factor 3 (IRF3) plays a crucial role in mediating cellular responses to virus intrusion. The protein kinase TBK1 is a key regulator inducing phosphorylation of IRF3. The regulatory mechanisms during IRF3 activation remain poorly characterized. In the present study, we have identified by yeast two-hybrid approach a specific interaction between IRF3 and chaperone heat-shock protein of 90 kDa (Hsp90). The C-terminal truncation mutant of Hsp90 is a strong dominant-negative inhibitor of IRF3 activation. Knockdown of endogenous Hsp90 by RNA interference attenuates IRF3 activation and its target gene expressions. Alternatively, Hsp90-specific inhibitor geldanamycin (GA) dramatically reduces expression of IRF3-regulated interferon-stimulated genes and abolishes the cytoplasm-to-nucleus translocation and DNA binding activity of IRF3 in Sendai virus-infected cells. Significantly, virus-induced IRF3 phosphorylation is blocked by GA, whereas GA does not affect the protein level of IRF3. In addition, TBK1 is found to be a client protein of Hsp90 in vivo. Treatment of 293 cells with GA interferes with the interaction of TBK1 and Hsp90, resulting in TBK1 destabilization and its subsequent proteasome-mediated degradation. Besides maintaining stability of TBK1, Hsp90 also forms a novel complex with TBK1 and IRF3, which brings TBK1 and IRF3 dynamically into proximity and facilitates signal transduction from TBK1 to IRF3. Our study uncovers an essential role of Hsp90 in the virus-induced activation of IRF3.

scholarly journals Structural and Functional Analysis of Interferon Regulatory Factor 3: Localization of the Transactivation and Autoinhibitory Domains

1999 ◽  
Vol 19 (4) ◽  
pp. 2465-2474 ◽  
Author(s):  
Rongtuan Lin ◽  
Yael Mamane ◽  
John Hiscott
Keyword(s):  
Amino Acids ◽  
Dna Binding ◽  
Virus Infection ◽  
Nuclear Translocation ◽  
Sendai Virus ◽  
Interferon Regulatory Factor ◽  
Phosphorylation Sites ◽  
Transactivation Domain ◽  
Regulatory Factor ◽  
Interferon Regulatory Factor 3

ABSTRACT The interferon regulatory factor 3 (IRF-3) gene encodes a 55-kDa protein which is expressed constitutively in all tissues. In unstimulated cells, IRF-3 is present in an inactive cytoplasmic form; following Sendai virus infection, IRF-3 is posttranslationally modified by protein phosphorylation at multiple serine and threonine residues located in the carboxy terminus. Virus-induced phosphorylation of IRF-3 leads to cytoplasmic to nuclear translocation of phosphorylated IRF-3, association with the transcriptional coactivator CBP/p300, and stimulation of DNA binding and transcriptional activities of virus-inducible genes. Using yeast and mammalian one-hybrid analysis, we now demonstrate that an extended, atypical transactivation domain is located in the C terminus of IRF-3 between amino acids (aa) 134 and 394. We also show that the C-terminal domain of IRF-3 located between aa 380 and 427 participates in the autoinhibition of IRF-3 activity via an intramolecular association with the N-terminal region between aa 98 and 240. After Sendai virus infection, an intermolecular association between IRF-3 proteins is detected, demonstrating a virus-dependent formation of IRF-3 homodimers; this interaction is also observed in the absence of virus infection with a constitutively activated form of IRF-3. Substitution of the C-terminal Ser-Thr phosphorylation sites with the phosphomimetic Asp in the region ISNSHPLSLTSDQ between amino acids 395 and 407 [IRF-3(5D)], but not the adjacent S385 and S386 residues, generates a constitutively activated DNA binding form of IRF-3. In contrast, substitution of S385 and S386 with either Ala or Asp inhibits both DNA binding and transactivation activities of the IRF-3(5D) protein. These studies thus define the transactivation domain of IRF-3, two domains that participate in the autoinhibition of IRF-3 activity, and the regulatory phosphorylation sites controlling IRF-3 dimer formation, DNA binding activity, and association with the CBP/p300 coactivator.

scholarly journals Loss of Interferon Regulatory Factor 3 in Cells Infected with Classical Swine Fever Virus Involves the N-Terminal Protease, Npro

2005 ◽  
Vol 79 (11) ◽  
pp. 7239-7247 ◽  
Author(s):  
S. Anna La Rocca ◽  
Rebecca J. Herbert ◽  
Helen Crooke ◽  
Trevor W. Drew ◽  
Thomas E. Wileman ◽  
...  
Keyword(s):  
Classical Swine Fever Virus ◽  
Viral Protein ◽  
Sindbis Virus ◽  
Classical Swine Fever ◽  
Interferon Regulatory Factor ◽  
Fever Virus ◽  
Luciferase Reporter ◽  
Regulatory Factor ◽  
Interferon Regulatory Factor 3 ◽  
Infected Cells

ABSTRACT We show that cells infected with the pestivirus classical swine fever virus (CSFV) fail to produce alpha/beta interferon not only following treatment with double-stranded RNA but also after superinfection with a heterologous virus, the alphavirus Sindbis virus, a virus shown to normally induce interferon. We investigated whether the inhibition of interferon synthesis by CSFV involved a block in interferon regulatory factor 3 (IRF3) activity. Cells infected with CSFV exhibited a lack of translocation of green fluorescent protein-IRF3 to the nucleus; however, constitutive shuttling of IRF3 was not blocked, since it could still accumulate in the nucleus in the presence of leptomycin B. Interestingly subcellular fractionation analysis showed that IRF3 was lost from the cytoplasm of infected cells from 18 h postinfection onwards. Using IRF3 promoter-luciferase reporter constructs, we demonstrate that loss of IRF3 was due to an inhibition of transcription of the IRF3 gene in CSFV-infected cells. Further, we investigated which viral protein may be responsible for the inhibition of interferon and loss of IRF3. We used cell lines expressing the CSFV N-terminal protease (Npro) to show that this single viral protein, unique to pestiviruses, inhibited interferon production in response to Sindbis virus. In addition to being lost from CSFV-infected cells, IRF3 was lost from Npro-expressing cells. The results demonstrate a novel viral evasion of innate host defenses, where interferon synthesis is prevented by inhibiting transcription of IRF3 in CSFV-infected cells.

scholarly journals Positive Regulation of Interferon Regulatory Factor 3 Activation by Herc5 via ISG15 Modification

2010 ◽  
Vol 30 (10) ◽  
pp. 2424-2436 ◽  
Author(s):  
He-Xin Shi ◽  
Kai Yang ◽  
Xing Liu ◽  
Xin-Yi Liu ◽  
Bo Wei ◽  
...  
Keyword(s):  
Sendai Virus ◽  
Specific Binding ◽  
Ectopic Expression ◽  
Interferon Regulatory Factor ◽  
Type I Interferons ◽  
Type I ◽  
Regulatory Factor ◽  
Interferon Regulatory Factor 3 ◽  
Antiviral Responses ◽  
Irf3 Activation

ABSTRACT Virus infection induces host antiviral responses, including induction of type I interferons. Transcription factor interferon regulatory factor 3 (IRF3) plays a pivotal role and is tightly regulated in this process. Here, we identify HERC5 (HECT domain and RLD 5) as a specific binding protein of IRF3 by immunoprecipitation. Ectopic expression or knockdown of HERC5 could, respectively, enhance or impair IRF3-mediated gene expression. Mechanistically, HERC5 catalyzes the conjugation of ubiquitin-like protein ISG15 onto IRF3 (Lys193, -360, and -366), thus attenuating the interaction between Pin1 and IRF3, resulting in sustained IRF3 activation. In contrast to results for wild-type IRF3, the mutant IRF3(K193,360,366R) interacts tightly with Pin1, is highly polyubiquitinated, and becomes less stable upon Sendai virus (SeV) infection. Consistently, host antiviral responses are obviously boosted or crippled in the presence or absence of HERC5, respectively. Collectively, this study characterizes HERC5 as a positive regulator of innate antiviral responses. It sustains IRF3 activation via a novel posttranslational modification, ISGylation.

scholarly journals Identification of TRAIL as an Interferon Regulatory Factor 3 Transcriptional Target

2005 ◽  
Vol 79 (14) ◽  
pp. 9320-9324 ◽  
Author(s):  
Jessica R. Kirshner ◽  
Alla Y. Karpova ◽  
Maren Kops ◽  
Peter M. Howley
Keyword(s):  
Viral Infection ◽  
Virus Production ◽  
Interferon Regulatory Factor ◽  
Progeny Virus ◽  
Regulatory Factor ◽  
Interferon Regulatory Factor 3 ◽  
Apoptosis Pathway ◽  
Infected Cells ◽  
Transcriptional Induction ◽  
Transcriptional Target

ABSTRACT Interferon production and apoptosis in virus-infected cells are necessary to prevent progeny virus production and to eliminate infected cells. Paramyxovirus infection induces apoptosis through interferon regulatory factor 3 (IRF-3), but the exact mechanism of how IRF-3 functions is unknown. We show that IRF-3 is involved in the transcriptional induction of TRAIL, a key player in the apoptosis pathway. IRF-3 upregulates TRAIL transcription following viral infection and binds an interferon-stimulated response element in the TRAIL promoter. The mRNA for TRAIL and its receptor, DR5, are induced following viral infection. These studies identify TRAIL as a novel IRF-3 transcriptional target.

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